Statement by
James F. Battey, Jr., M.D., Ph.D.
National Institute on Deafness and Other Communication Disorders, and
NIH Stem Cell Task Force
National Institutes of Health
U.S. Department of Health and Human Services

Research on Alternative Sources of Pluripotent Stem Cells

The Subcommittee on Labor, Health and Human Services, Education, and Related Agenices
Committee on Appropriations
United States Senate

Tuesday, June 27, 2006

Mr. Chairman, Senator Harkin, and Members of the Subcommittee, I am pleased to appear before you today to testify about stem cell research. As you are aware, I previously testified to this Subcommittee about human embryonic stem cells (hESC) as a tool for advancing our knowledge about cell specialization, and its great potential to be medically valuable. However, using established methods, these cannot be obtained without destroying human embryos. There have been recent publications about alternative ways to establish human pluripotent stem cell lines that claim to avoid the issue of creating, destroying, or harming human embryos. In 2005, the President's Council on Bioethics published a white paper on "Alternative Sources of Human Pluripotent Stem Cells." My testimony will provide some information on the scientific advances highlighted in that report.

Pluripotent Stem Cells from Nonviable Embryos
Scientists proposing this method noted that during the human in vitro fertilization (IVF) process, there are numerous embryos that fail to continue to divide and are therefore judged to be unsuitable for implantation.

Recently, in a privately funded study, the scientists evaluated the physical characteristics of human embryos created for IVF but not used because they were considered to be "nonviable." The scientists observed that many of the nonviable embryos had fewer cells than normal, and failed to compact into a morula or a blastocyst, which are developmental stages of the embryo. They propose that nonviable embryos with these features of arrested development at 5 days post-feritilization be considered "dead." This would allow scientists to harvest cells from nonviable human embryos in experimental efforts to generate human embryonic stem cell lines. (Regen. Med. 1: 367-371, D.W. Landry, H.A. Zucker, M.V. Sauer, M. Reznik, L. Wiebe).

To date, there is no published study showing that it is possible to generate an embryonic stem cell line from a non-dividing embryo in rodents, non-human primates, or humans. If stem cell lines could be derived from such embryos, the resulting cell line would have to be carefully monitored for karyotypic (genetic) abnormalities or other defects.

Pluripotent Stem Cells from Biopsied Blastomeres
This proposal involves creating an embryonic stem cell line by using a blastomere cell from an embryo. When performing pre-implantation genetic diagnosis (PGD), a single blastomere cell is removed from an 8-cell stage embryo (approximately Day 3 in embryo development where all cells are assumed to be totipotent) for genetic analysis, and the remaining seven cells constituting the embryo are used for reproductive purposes through the standard IVF procedure. The proposal states that a single cell, or several cells, might be removed from an embryo at the 8-cell stage at the same time the embryo is undergoing PGD, and these additional cell(s) could be used for the purpose of creating a hESC line.

Recently, privately funded scientists removed (i.e., biopsied) single cells from early mouse embryos and used them to establish mouse embryonic stem cell lines. The remaining cells of the embryo were implanted in surrogate mouse wombs, and approximately half developed into seemingly normal mouse pups. In the control group of embryos that did not undergo biopsies, about half also developed to birth as normal pups. This research is the first to demonstrate that single cell embryo biopsy can be used successfully to generate stem cell lines. If this technique succeeds with human embryos, it may provide another way to generate human embryonic stem cell lines. Although single cell embryo biopsy proposes to avoid embryo destruction, scientists do not yet know how much risk the procedure might confer to an otherwise healthy human embryo. (Nature 439:216-219, laboratory of R. Lanza)

NIH believes that such experiments could and should be pursued in non-human primates. If this approach is successful, the resulting stem cell lines would, of course, have to be validated for genetic stability, pluripotency, and unlimited self-renewal - all cardinal features of embryonic stem cell lines generated from blastocysts by culturing the inner cell mass.

Pluripotent Stem Cells from Biological Artifacts
Proponents of this method assert that it may be possible to do the following: (1) genetically modify a somatic cell in culture, for instance, the cell might be engineered to lack a gene or genes crucial for cell-to-cell signaling or the integrated organization essential for normal embryogenesis; (2) use this genetically modified somatic cell as the source of a nucleus and genome for somatic cell nuclear transfer (SCNT) into a human oocyte. This method is referred to as Altered Nuclear Transfer (ANT); (3) allow the resulting entity to develop to a point when it may yield embryonic-like stem cells; and (4) after extraction, attempt to generate a hESC or hESC-like line from these cells.

ANT is a general concept that its proponents suggest could take a number of specific forms. One version of the idea proposes that scientists turn off a gene needed for implantation in the uterus (Cdx2) in the patient cell nucleus before it is transferred into the donor egg. NIH-supported scientists recently reported proof of principle tests that ANT works in mice. Mouse ANT entities whose Cdx2 gene is switched off are unable to implant in the uterus and do not survive to birth. However, scientists used ANT to create viable stem cell lines capable of producing almost all cell types. The scientists point out that this technique must still be tested with monkey and human donor nuclei, and the manipulation needed to control Cdx2 expression introduces another logistical hurdle that may complicate ANT's use to derive embryonic stem cells. (Nature 439(7073):212-5, laboratory of R. Jaenisch)

Pluripotent Stem Cells by Reprogramming Somatic Cells
This proposal involves reprogramming human somatic cells, perhaps with the aid of special cytoplasmic factors obtained from oocytes (or from pluripotent embryonic stem cells), so as to "dedifferentiate" them back into pluripotent stem cells. Crucial to this approach is discovering a way to reverse cell differentiation all the way back to pluripotency, but not further back to totipotency.

Scientists in Germany recently succeeded in coaxing adult mouse stem cells that normally produce sperm (spermatogonial stem cells, or SSCs) to instead behave in a manner similar to embryonic stem cells. They accomplished this switch of fate by finding the elusive SSCs in mouse testicles and growing them in the laboratory under standard embryonic stem cell culture conditions. Under those conditions, the cells made several proteins characteristic of embryonic stem cells. The scientists subjected the cells to critical tests for pluripotency, and their results suggest that the cells can become any type of cell in the body. As a result, the scientists named them multipotent adult germline stem cells (maGSCs). If scientists can find similar cells in human testicles, the cells could provide a new source of patient-specific stem cells, and could also provide more pluripotent cell lines for research. (Nature advance online publication, laboratory of G. Hasenfuss)

In another study, privately funded scientists fused cultured adult human skin cells with hESCs. The resulting "hybrid" cells had many characteristics of hESCs-they grew and divided in a similar manner and manufactured proteins that are typically made in hESCs. Some as-yet unknown factor(s) within the hESCs enabled them to "reprogram" the adult skin cells to behave as hESCs. The cells still raise a significant technical barrier that must be overcome before they can be used to treat patients. Because fused cells are tetraploid (i.e., they contain four copies of the cellular DNA rather than the normal two copies), scientists must develop a method to remove the extra DNA without eliminating their hESC-like properties. If this hurdle can be overcome, this technique may one day allow scientists to create patient-specific stem cells without using human eggs. At present, this new approach to creating stem cells is a useful model system for studying how stem cells "reprogram" adult cells to have properties of pluripotent cells. (Science 309:1369-1373, laboratory of K. Eggan)

Privately funded scientists in the United Kingdom now report that the reprogramming process in mice is more efficient when they engineer the stem cells to over-express Nanog, a gene important for maintaining stem cells' self-renewing properties. The scientists reported a 200-fold increase in the efficiency of the process when mouse embryonic stem cells that over-expressed Nanog were fused with stem cells from mouse brain; however, the fused cells are tetraploid. This study demonstrates that Nanog plays an important role in reprogramming the mouse brain cells to a state of pluripotency. If these results can be repeated with human cells, they would represent a first step toward learning how to reprogram adult cells to behave as stem cells and directing them to become specific cell types for use in treating human beings. (Nature Advance Online Publication 14 June 2006; lab of A. Smith)

NIH welcomes the receipt of investigator-initiated grant applications on these research topics. As with all grant applications, such proposals would be judged for scientific merit by peer review. We are very grateful for your continued support. I will be happy to try to answer any questions that you might have.

Last Revised: June 28, 2006